The high affinity receptor for IgE on mast cells and basophils (Fc epsilon RI) plays a central role in immediate hypersensitivity reactions. Aggregation of receptor-bound IgE by polyvalent antigen leads to aggregation of the receptors and cellular secretion of both performed and newly synthesized mediators of inflammation. The molecular mechanisms by which aggregation of the receptors generate these cellular responses are the central focus of these studies. During the past year we continued to employ mutated receptors in order to establish which portions of the receptor participate in its principal functions. Our new results show: 1) No single cytoplasmic domain is required for receptors to localize to coated pits and thereafter to become internalized after aggregation. However, the lipid-anchored ectodomain of the alpha subunit fails to participate in this process. 2) Previous results with transfected P815 mastocytoma cell suggested a critical role for the gamma subunit. Because the effects on signaling by the endogenous Fc receptors for IgG closely paralled the effects on Fc epsilon RI, we postulated that of the three types of Fc receptors for IgG on these cells, only type III would be capable on initiating a variety of biochemical changes. This was confirmed by studying rat mucosal mast cell tumor line (RBL) transfected with individual Fc receptor isoforms. 3) One of the earliest consequences of receptor aggregation (but not of stimulation of the cells with phorbol esters) is phosphorylation of tyrosines on the beta and gamma subunits of the receptor. We have been successful in retaining some of this activity on broken cell preparations of RBL cells. This should greatly assist our efforts to elucidate the molecular events that result from aggregation of the receptor.